EFFECTS OF ENERGY SOURCES ON MILK PRODUCTION AND REPRODUCTION OF DAIRY COWS

Transcription

1 EFFECTS OF ENERGY SOURCES ON MILK PRODUCTION AND REPRODUCTION OF DAIRY COWS By Bilungi Alain Useni Dissertation presented for the degree of Doctor of Philosophy in Animal Science in the Faculty of AgriScience at Stellenbosch University Promotor: Dr C.J.C. Muller Co-Promotor: Prof C.W. Cruywagen December 2017

2 DECLARATION By submitting this dissertation electronically, I declare that the entirety of the work contained therein is my own, original work, that I am the authorship owner thereof (save to the extent explicitly otherwise stated), that reproduction and publication thereof by Stellenbosch University will not infringe any third party rights and that I have not previously in its entirety or in part submitted it for obtaining any qualifications. DATE: December 2017 Bilungi Alain Useni Copyright 2017 Stellenbosch University All rights reserved ii

3 ABSTRACT Two main objectives were investigated in this study, conducted at the Elsenburg Research Farm of the Western Cape Department of Agriculture (WCDA). The first objective was to study the effect of nutritional treatments differing after calving in energy levels and sources on plasma metabolites and live weight (LW) changes, milk production and milk composition, ovarian follicular dynamics and oestrous activity, as well as fertility traits of dairy cows. The second objective was to investigate the effects of two late prepartum diets differing in non-fibre carbohydrates (NFC) and protein levels on milk production and milk composition, LW changes and fertility traits of dairy cows in the subsequent lactation. Three trials were conducted in this study. The first trial explored the effect of energy-dense ingredients like fat or starch on milk production and milk composition, plasma metabolites, LW change and fertility of lactating dairy cows. In this study, pregnant Holstein heifers and dry cows were similarly managed and fed prepartum. Following calving, they were allowed to graze ad libitum on kikuyu-ryegrass pasture, and received supplements that differed in terms of inclusion levels and types of concentrates. Holstein cows in the control group received 7 kg/day of a control concentrate supplement for both primiparous and multiparous groups, while treatments consisted of 11.6 and 12.6 kg/day for primiparous and multiparous cows, respectively. Supplements in treatments contained high starch-low fat (HSLF) and high starch-low fat/low starch-high fat (HSLF-LSHF) levels. The control and HSLF concentrates were offered from calving to 154 days in milk (DIM). The HSLF supplement was a glucogenic concentrate containing maize as the energy source. The HSLF-LSHF supplements were combinations of a glucogenic concentrate offered for the first 60 DIM as per treatment HSLF and followed from 61 to 154 DIM by a lipogenic concentrate containing wheat bran and calcium (Ca)-salts of long-chain fatty acids as the energy sources. The postpartum diets were formulated to be isonitrogenous. The control diet contained low energy level (2.47 Mcal ME/kg DM) and provided 457, 104 and 40 g/kg of NDF, starch and fat on DM basis, respectively. The HSLF diet contained high energy level (2.71 Mcal ME/kg DM) and offered 341, 242 and 35 g/kg of NDF, starch and fat on DM basis, respectively. The HSLF treatment was formulated to increase plasma insulin and glucose levels in order to reduce the magnitude of the NEB and encourage the early return of oestrous cyclicity after calving. In the HSLF-LSHF combination treatment, the high starch-based diet initially aimed to achieve the same objective of the HSLF treatment during the first 60 DIM. The LSHF diet contained high energy level (2.70 Mcal ME/kg DM) and provided 388, 137 and 58 g/kg of NDF, starch and fat on DM basis, respectively. Secondly, the LSHF diet was fed from 61 DIM to decrease plasma insulin and iii

4 improve plasma cholesterol in order to increase the size of the ovulatory follicle as well as promote embryo development. The second experiment trial was simultaneously conducted with the first trial using the same feeding treatments. In this trial, the same objectives of the diets were evaluated for ovarian follicular dynamics and oestrous activity of multiparous cows after being synchronized by means of the Ovsynch protocol at 80 ± 10 DIM without artificial insemination (AI). Results on plasma metabolites showed that postpartum levels of non-esterified fatty acids (NEFA) and β- hydroxybutyrate (BHB) did not differ between nutritional treatments for multiparous cows, but levels increased (P < 0.05) for the control in comparison to HSLF and HSLF-LSHF treatments for primiparous animals. In addition, plasma urea levels were higher (P < 0.05) in both primiparous and multiparous animals receiving HSLF and HSLF-LSHF concentrates, compared to the control concentrate. The postpartum LW was the lowest (P < 0.05) and LW loss was greater (P < 0.05) in cows in the control in comparison to HSLF and HSLF-LSHF treatments in both primiparous and multiparous cows. Findings on productive responses showed that the milk yields and 4 % fat corrected milk (FCM) yields of both primiparous and multiparous cows in HSLF and HSLF-HFLS treatments were higher (P < 0.05), compared to cows in the control. In the primiparous group, the HSLF diet enhanced (P < 0.05) milk protein content whereas HSLF and HSLF-HFLS diets resulted in increased milk lactose content (P < 0.05). In multiparous group, HSLF treatment increased (P < 0.05) milk lactose content. Results of ovarian follicular dynamics and oestrous activity showed that dimensions of the ovaries and the preovulatory follicle as well as numbers of follicles within different classes in dairy cows were similar between nutritional treatments. However grazing cows receiving high (12.6 kg/day) levels of concentrates in the HSLF and HSLF-LSHF treatments recorded a higher number of total follicles (P < 0.05), compared to those on the low (7 kg/day) level of concentrate in the control group. Results of reproductive performance showed no nutritional effects in both primiparous and multiparous cows on the interval calving to first service, the percentage of cows serviced for the first time within the 80 DIM, pregnancy rate at first service, number of services per conception, interval days open and the pregnancy rate at 100 DIM. However, the conception rate at 150 DIM in both primiparous and multiparous groups improved (P < 0.05) in cows fed HSLF and HSLF-LSHF concentrates, in comparison to the control. The third trial was conducted to evaluate the effect of two late prepartum nutritional treatments on subsequent LW changes, milk production and milk composition, and fertility traits of Holstein dairy cows. Pregnant heifers and dry cows received a similar level and type of prepartum concentrate iv

5 associated either unchopped oat hay ad libitum (control) or ad libitum intake of a partial total mixed ration (ptmr, treatment). The ptmr consisted of oat hay (48%), lucerne hay (43%) and soybean oil cake meal (9%). The prepartum treatment was designed to optimise rumen digestion and fermentation through improved prepartum intake of NFC and protein per kg on DM basis with an objective of reducing the extent of a periparturient negative energy balance (NEB) in comparison to the control. Following parturition, primiparous and multiparous cows received 7 kg/day of concentrate while grazing ad libitum on kikuyu-ryegrass pastures from calving to 120 DIM. Results showed that prepartum LW was similar for both primiparous and multiparous groups. In the primiparous group, the prepartum treatment significantly improved (P < 0.05) postpartum LW, with 20 kg more weight at LW nadir, compared to the control. However, no prepartum nutritional effects were detected on postpartum LW traits and LW loss for multiparous cows. Milk yields and milk composition were similar between the control and treatment for both primiparous and multiparous groups. Primiparous cows in the control group had a longer interval from calving to first service (P < 0.05), compared to those in treatment group. However, this improvement did not turn into increased conception rates of primiparous cows at 120 DIM. Similarly, prepartum diet effects in multiparous cows showed no postpartum differences in conception rates at 120 DIM. It can concluded that there is definite merit in the use of energy levels and sources for improving production responses, plasma metabolite profiles and LW change, ovarian follicular dynamics and fertility of grazing dairy cows. These improvements are related to nutrient composition and levels of total energy intake, controlling the profile of nutrients absorbed in the gastrointestinal tract (GIT) and their hepatic partitioning to various body tissues for metabolic requirements of dairy cows. This modulation has probably influenced the hypothalamic-pituitary-ovarian-uterine axis through improved nutritional status and optimized the energy carry-over effect into fertility success, while sustaining milk production. However, feeding fibre-based diets differing in NFC and protein levels in the late prepartum period showed no benefit in either milk production or conception rates of dairy cows in the subsequent lactation. The lack of significant differences of prepartum dietary effects on milk production and conception success can be related to the prepartum feeding period and the relatively low number of animals per treatment used in the study. Further investigations should be undertaken using different inclusion and types of energy sources in prepartum and postpartum dairy cows on metabolic and hormonal responses that can improve restoration body reserves and benefit chronological sequences of the reproductive process such as postpartum health, uterine regression, resumption of oestrous cycles, conception, and embryo/pregnancy survival until the next calving. v

6 OPSOMMING Hierdie studie is uitgevoer op die Elsenburg Navorsingsplaas van die Wes-Kaapse Departement van Landbou (WKDL) en het twee hoofdoelwitte gehad. Die eerste doelwit was om die effek te bepaal van verskilllende voedingspeile, naamlik twee energiepeile en twee energiebronne, op spesifieke bloedplasma metaboliete, die verandering in liggaamsmassa, melkproduksie en -samestelling, ovarium en follikulêre aktiwiteite asook vrugbaarheidseienskappe van lakterende Holsteinkoeie wat die diete vanaf na kalwing ontvang het. Die tweede doelwit was om die effek te bepaal van twee diete, verskillend ten opsigte van nie-vesel koolhidraat- (NVK) en proteïenpeile, wat tydens die laat-dragtige periode aan melkkoeie gevoer is op die melkproduksie en -samestelling, verandering in liggaamsmassa en vrugbaarheidseienskappe van melkkoeie in die daaropvolgende laktasie. Vir die eerste doelwit is drie studies uitgevoer. In die eerste studie is die effek van energieryke grondstowwe soos vet of stysel op die melkproduksie en -samestelling, plasma metaboliete, liggaamsmassaverandering en die vrugbaarheid van lakterende melkkoeie bepaal. In hierdie studie is dragtige Holsteinverse en droë koeie voor kalwing op dieselfde manier bestuur en gevoer. Na kalwing het koeie ad libitum toegang gehad tot aangeplante kikoejoe-raaigrasweiding met kragvoeraanvullings wat verskil het ten opsigte van insluitingspeil en die tipe energiebron. Die kontrole-dieet het bestaan uit n lae kragvoerpeil (7 kg/koei/dag) van n standaard aanvulling terwyl behandelings bestaan het uit aanvullings wat (1) hoë stysel en lae vetpeile (HSLV) en (2) hoë stysel en lae vetpeile gevolg deur lae stysel en hoë vetpeile (HSLV-LSHV) bevat het. Dié aanvullings is teen hoër peile (11.6 en 12.4 kg/koei/dag vir eerste- en tweede-pluslaktasiekoeie, onerskeidelik) as die standaard kragvoer aanvulling voorsien. Die HSLV-aanvulling het bestaan uit n konsentraat wat glukose bevat met mielies as energiebron en is vanaf kalwing tot 154 dae na kalf aan koeie voorsien. Die aanvullings wat in die behandeling HSLV-LSHV gebruik is, het bestaan uit kombinasies van n glukose-bevattende energiebron vir die eerste 60 dae van die laktasieperiode, soortgelyk aan die behandeling HSLV, gevolg deur n kragvoer (LSHV) wat vet bevat het as energiebron (koringsemels en kalsiumsoute met langkettingvetsure) wat vanaf 61 tot 154 dae na kalf aan koeie gevoer is. Die tweede proef is gesamentlik met die eerste proef gedoen met dieselfde diete. Koeie wat op 80 ± 10 dae na kalf nog nie kunsmatig geïnsemineer was nie, is gesinchroniseer volgens die Ovsynch program en die follikulêre veranderinge en hitte-aktiwiteit van koeie is bepaal. Die energiebalansstudie het aangetoon dat geen verskille voorgekom het ten opsigte van die nie-esterbevattende vetsure (NEVS), plasma ureumpeile en liggaamsmassaveranderinge vir beide eerste- en vi

7 tweedeplus-laktasiekoeie nie. Die NEVS en β-hidroksiebutiraat (BHB) peile na-kalwing is ook nie beïnvloed vir die tweede-pluslaktasiekoeie nie terwyl die peile hoër (P < 0.05) was by eerstelaktasiekoeie wat die kontrole diet ontvang het in vergelyking met koeie wat die HSLV en HSLV-LSHV aanvullings ontvang het. Ureumpeile in bloedplasma in beide die eerste- en tweedeplus-laktasiekoeie is eweneens beïnvloed (P < 0.05) deur diëte met laer peile wat die kontrole dieet in vergelyking wat die HSLV en HSLV-LSHV aanvullings ontvang het. Aanvullend hierby het beide die eerste- en tweedeplus-laktasiekoeie wat die kontrole aanvulling ontvang het n groter afname in liggaamsmassa ondervind in vergelyking met koeie wat die HSLV en HSLV- LSHV aanvullings ontvang het. Die produksiestudie het getoon dat die melkproduksie van koeie verhoog (P < 0.05) en dat sommige bestanddele in melk ook verander (P < 0.05) is deur die behandelings wat toegepas is. Die melkproduksie van beide eerstelaktasie- en ouer (tweedepluslaktasie) koeie wat die HSLV en HSLV-LSHV behandelings ontvang het, was hoër as die melkproduksie van koeie wat die kontrole dieet ontvang het. Eerstelaktasiekoeie wat die HSLV aanvulling ontvang het, het melk met n hoër (P < 0.05) proteienpersentasie geproduseer terwyl die melk van dié koeie wat die HSLV-LSHV aanvulling ontvang het, n hoër (P < 0.05) laktosepersentasie bevat het. Netso het die melk van tweede-pluslaktasiekoeie wat die HSLV aanvulling ontvang het, hoër laktose peile bevat. Die follikulêre aktiwiteite in die ovaria en geslagsaktiwiteit is nie beïnvloed deur die vlak van energie-inname nie terwyl die totale aantal follikels wel beïnvloed (P < 0.05) is. Die reproduksieprestasie van beide eerste- en tweedepluslaktasiekoeie vir die volgende reproduksie-eienskappe, naamlik periode vanaf kalf tot eerste inseminasie, die proporsie van koeie gedek voor 80 dae-in-melk, die persentasie besetting met eerste inseminasie, aantal dekkings per konsepsie, die aantal dae oop en die proporsie koeie wat beset was by 100 dae-in-melk, is nie beïnvloed deur kragvoeraanvullings nie. Die proporsie koeie wat beset was by 150 dae-in-melk is wel beïnvloed (P < 0.05) deur kragvoeraanvulling met meer eerstelaktasie- en tweedeplus-laktasie koeie beset wat die HSLV en HSLV-LSHV aanvullings ontvang het in vergelyking met die kontrole-aanvulling. Die derde studie is uitgevoer om die invloed van die energiepeil in die dieet voor kalwing ten opsigte van liggaamsmassaverandering, melkproduksie en vrugbaarheidseienskappe gedurende die daaropvolgende laktasieperiode te bepaal. Dragtige verse en droë koeie is voor kalwing dieselfde tipe en hoeveelheid kragvoer voorsien tesame met (1) ad libitum gevoerde lang, ongekerfde hawerhooi en (2) n ad libitum inname van n gedeeltelike totaal gemengde rantsoen (ptmr) wat uit 48% hawerhooi, 43% lusernhooi en 9% sojaboon-oliekoekmeel bestaan het. Na kalwing is die eerstelaktasie- en tweedeplus-laktasiekoeie op aangeplante kikoejoe-raaigrasweiding aangehou en vii

8 het hulle dieselfde kragvoeraanvulling ontvang. Volgens die proefresultate het die voorkalwingsliggaamsmassa van beide eerstelaktasie- en tweedeplus-laktasiekoeie in die verskillende behandelingsgroepe nie verskil nie. Hierteenoor het die na-kalwingsliggaamsmassa van eerstelaktasiekoeie wat die kontrole-dieet (hawerhooi) en ptmr-voorkalwingsdieet ontvang het, verskil terwyl geen effek waargeneem is by die tweedeplus-laktasiekoeie nie. Die melkproduksie en samestelling was dieselfde vir beide eerstelaktasie- en tweedeplus-laktasiekoeie wat die kontrole en behandelingsdiëte voorkalwing ontvang het. Eerstelaktasiekoeie wat net hawerhooi gedurende die voorkalwingsperiode ontvang het, het n langer (P < 0.05) periode van kalf tot eerste inseminasie gehad in vergelyking met koeie wat die ptmr-dieet ontvang het. Geen verskille is gevind vir tweedeplus-laktasiekoeie nie. Net so is geen verskille tussen die kontrole en proefdiete gevind by beide eerstelaktasie- en tweedeplus-laktasiekoeie vir die persentasie koeie wat vir die eerste keer gedek is binne 80 dae na kalf, dragtigheid na eerste inseminasie, koeiliggaamsmassa met eerste dekking en besettingspersentasie teen 120 dae-in-melk. Resultate toon dat daar beslis meriete bestaan om spesifieke energiebronne te gebruik om die volgende eienskappe te verbeter, naamlik melkproduksie, status van die enegiebalans en verandering in die liggaamsmassa van koeie asook follikulêre aktiwiteite, wys van hitte en die vrugbaarheid van koeie. Hierdie verbeteringe hou verband met voedingstofsamestelling en totale energie-innamevlakke, wat die profiel van voedingstowwe wat in die spysverteringskanaal (SVK) geabsorbeer word, sowel as die allokasie daarvan in die lewer na die onderskeie liggaamsweefsels vir die metaboliese behoeftes van melkkoeie, beheer. Die verandering in voedingstowwe het waarskynlik die hypothalamus-pituitêre-ovarium verbintenis beïnvloed tot n beter energie balans met n verdere oordragingseffek op die vrugbaarheid van koeie terwyl melkproduksiepeile gehandhaaf word. Tersefdertyd het n verandering in die dieet wat dragtige verse en droëkoeie voorkalwing ontvang het geen voordele ingehou vir koeie in die daaropvolgende laktasieperiode ten opsigte van melkproduksie, -samestelling of besettingspersentasies nie. Dit kan waarskynlik toegeskryf word aan die beperkte aantal koeie wat in die proef in elke behandeling gebruik is. In hierdie studie het die behandelings nie-betekenisvolle produksie- en reproduksieresponse op alle gebiede getoon nie. Dit word aanbeveel dat die studie opgevolg behoort te word met groter getalle diere. viii

9 ACKNOWLEDGEMENTS On the completion of this work, I would like to express my sincerest appreciation and gratitude to the following people, without whom this work would have been impossible: My promotor, Dr C.J.C. Muller, for his dedication, guidance, patience and endless support during my studies. This achievement would not have been possible without your contribution and support, I respect and salute you for that; Prof. C.W. Cruywagen, for academic support towards my person and for agreeing to co-promote this study. From my heart, I thank you for empowering me through education; The Western Cape Department of Agriculture at Elsenburg for allowing me to conduct the study and for the use of their facilities; The Dairy Section of the Western Cape Department of Agriculture at Elsenburg, for the workforce support during the trials of this study; The Trustees of the Western Cape Agricultural Research Trust for funding the study; The Stellenbosch Veterinary Hospital, Dr. A. Kidd, for all veterinary assistance and ultrasound procedures; The Dairy Laboratory of the Agricultural Research Council at Elsenburg, Stellenbosch, for the analyses of the milk samples; The Durbanville Vet Diagnostics Laboratory, for the analysis of plasma biomarkers; Nova Feeds, Malmesbury for manufacturing the experimental concentrates according to project requirements The Durbanville MouldProd Ltd., for the assistance and training in handling the ultrasound scanner; Dr Rachel M. Isango Useni, my lovely wife, for her patience and endless support during my studies. For all of that and for much more you do for me and the family, I say I thank you, I m blessed to have you and I dearly love you; Shukrani A. Joshua Useni, my son and big friend, for his cheerful smiles and always playing at my feet at first and trying to assist me in his ways as his age progresses when writing this document; Neema W. Amelia Useni, my little angel, for much joy and blessings that you bring to the family; My parents, brothers and sisters, for their supports and encouragements in life; ix

10 My friends and comrades in the field of practice, for continued encouragements; Most of all, Jesus Christ, for making it possible through Him. Much thanks! Enkosi kakhulu! Baie dankie! Merci beaucoup! Asante sana! x

11 DEDICATION J'ai encore vu sous le soleil que la course n'est point aux agiles ni la guerre aux vaillants, ni le pain aux sages, ni la richesse aux intelligents, ni la faveur aux savants; car tout dépend pour eux du temps et des circonstances. Ecclésiaste 9: 11 To my wife Rachel, my son Joshua and my daughter Amelia To whom my success and progress are their main concerns, To whom, by grace, makes the impossible possible, I dedicate this work. xi

12 NOTES The language and style used in this thesis are in accordance with the requirements of the South African Journal of Animal Science. This thesis represents a compilation of manuscripts where each chapter is an individual entity and some repetition between chapters has been unavoidable. xii

13 CONFERENCE CONTRIBUTIONS Oral presentations Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of energy sources on the milk production and reproduction of lactating Holstein cows. 6th All Africa Conference on Animal Agriculture, Nairobi, Kenya, October Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of energy sources on energy balance & milk production of dairy cows. 48 th SASAS Congress, 48 th SASAS Congress, Empagneni. South Africa, September, Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of different feeding regimes during the dry period on subsequent live weight changes, milk production and fertility traits of dairy cows. 49 th SASAS Congress, Spier, Stellenbosch, South Africa, 3-6 July Poster presentations Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of feeding management during transition on energy balance and early lactation of dairy cows. 47 th SASAS Congress, University of Pretoria, Pretoria, South Africa. 6-8 July Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of different levels and types of energy sources on some fertility traits of Holstein cows. 48 th SASAS Congress, 48 th SASAS Congress, Empagneni, South Africa, September, Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of energy sources on ovarian follicular dynamics and oestrous activity of dairy cows. 49 th SASAS Congress, Spier, Stellenbosch, South Africa, 3-6 July Publication Useni, B.A., Muller, C.J.C. & Cruywagen, C.W., Effect of energy sources on the milk production and reproduction of lactating Holstein cows. East Afr. Agric. Forest. J. 81, xiii

14 TABLE OF CONTENTS DECLARATION... ii ABSTRACT... iii OPSOMMING... vi ACKNOWLEDGEMENTS... ix DEDICATION... xi NOTES... xii CONFERENCE CONTRIBUTIONS... xiii TABLE OF CONTENTS... xiv LISTS OF FIGURES... xvii LIST OF TABLES... xx LIST OF ABBREVIATIONS... xxi Chapter General introduction References... 6 Chapter Literature review Introduction Energy sources during the dry period Negative energy balance and postpartum related disorders in dairy cows Energy partitioning in dairy cows Effect of energy sources on metabolism of dairy cows Effect of energy sources on milk yield and milk composition Effect of energy sources on reproduction efficiency of dairy cows Conclusion References Chapter Effect of energy sources on blood metabolite profiles and live weight changes as an indication of the nutritional status in Holstein cows Abstract Introduction Materials and Methods xiv

15 Experimental location Experimental husbandry, animals and diets Experimental sampling and data collection Statistical analysis Results and Discussion Conclusion References Chapter Effect of energy sources on the milk production and milk composition of Holstein cows Abstract Introduction Materials and Methods Experimental location Experimental husbandry, animals and diets Experimental sampling and data collection Statistical analysis Results and Discussion Conclusion References Chapter Effect of energy sources on ovarian follicular dynamics and oestrous activity of Holstein cows Abstract Introduction Materials and Methods Experimental location Experimental husbandry, animals and diets Experimental sampling and data collection Statistical analysis Results and Discussion Conclusion References xv

16 Chapter Effect of different energy levels and sources on the reproductive performance of Holstein cows Abstract Introduction Materials and Methods Experimental location Experimental husbandry, animals and diets Experimental sampling and data collection Statistical analysis Results and Discussion Conclusion References Chapter The effect of late prepartum feeding programmes on the live weight changes, milk production and fertility traits in the subsequent lactation period of Holstein cows Abstract Introduction Materials and Methods Experimental location Experimental husbandry, animals and diets Experimental sampling and data collection Statistical analysis Results and Discussion Conclusion References Chapter General conclusions and Recommendations xvi

17 LISTS OF FIGURES Figure 3.1 A graphic representation of pasture quality parameters monitored over the duration of the study. Figure 3.2 The effect of nutritional treatments differing after calving in energy levels and sources on non-esterified fatty acid (mean ± SE) levels recorded for primiparous dairy cows during a four week prepartum to 13 week postpartum period. Figure 3.3 The effect of nutritional treatments differing after calving in energy levels and sources on the non-esterified fatty acid (mean ± SE) levels recorded for multiparous dairy cows during a four week prepartum to 13 week postpartum period. Figure 3.4 The effect of nutritional treatments differing after calving in energy levels and sources on the β-hydroxybutyrate (mean ± SE) levels recorded for primiparous dairy cows during a 13 week postpartum period. Figure 3.5 The effect of nutritional treatments differing after calving in energy levels and sources on the β-hydroxybutyrate (mean ± SE) levels recorded for multiparous dairy cows during a four week prepartum to 13 week postpartum period. Figure 3.6 The effect of nutritional treatments differing after calving in energy levels and sources on the plasma urea (mean ± SE) levels recorded for primiparous dairy cows during a four week prepartum to 13 week postpartum period. Figure 3.7 The effect of nutritional treatments differing after calving in energy levels and sources on the plasma urea (mean ± SE) levels recorded for multiparous dairy cows during a four week prepartum to 13 week postpartum period. Figure 3.8 The effect of nutritional treatments differing after calving in energy levels and sources on the live weight (mean ± SE) recorded for primiparous dairy cows during an eight week prepartum to 22 week postpartum period. Figure 3.9 The effect of nutritional treatments differing after calving in energy levels and sources on the postpartum live weight loss (mean ± SE) recorded for primiparous dairy cows during a 22 week postpartum period. Figure 3.10 The effect of nutritional treatments differing after calving in energy levels and sources on the live weight (mean ± SE) recorded for multiparous dairy cows during an eight week prepartum to 22 week postpartum period. Figure 3.11 The effect of nutritional treatments differing after calving in energy levels and sources on the postpartum live weight loss (mean ± SE) recorded for multiparous dairy cows during a 22 week postpartum period. xvii

18 Figure 4.1 The effect of nutritional treatments differing after calving in energy levels and sources on the milk yield (mean ± SE) recorded for primiparous dairy cows during a 22 week postpartum period. Figure 4.2 The effect of nutritional treatments differing after calving in energy levels and sources on the 4% fat corrected milk (FCM) yield (mean ± SE) recorded for primiparous dairy cows during a 22 week postpartum period. Figure 4.3 The effect of nutritional treatments differing after calving in energy levels and sources on the milk yield (mean ± SE) recorded for multiparous dairy cows during a 22 week postpartum period. Figure 4.4 The effect of nutritional treatments differing after calving in energy levels and sources on the 4% fat corrected milk (FCM) yield (mean ± SE) recorded for multiparous dairy cows during a 22 week postpartum period. Figure 4.5 The effect of nutritional treatments differing after calving in energy levels and sources on the milk lactose content (mean ± SE) recorded for primiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.6 The effect of nutritional treatments differing after calving in energy levels and sources on the milk lactose content (mean ± SE) recorded for multiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.7 The effect of nutritional treatments differing after calving in energy levels and sources on the milk fat content (mean ± SE) recorded for primiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.8 The effect of nutritional treatments differing after calving in energy levels and sources on the milk fat content (mean ± SE) recorded for multiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.9 The effect of nutritional treatments differing after calving in energy levels and sources on the milk protein content (mean ± SE) recorded for primiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.10 The effect of nutritional treatments differing after calving in energy levels and sources on the milk protein content (mean ± SE) recorded for multiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 4.11 The effect of nutritional treatments differing after calving in energy levels and sources on the milk urea nitrogen (MUN) levels (mean ± SE) recorded for primiparous dairy cows at a 35 days interval during a 154 day postpartum period. xviii

19 Figure 4.12 The effect of nutritional treatments differing after calving in energy levels and sources on the milk urea nitrogen (MUN) levels (mean ± SE) recorded for multiparous dairy cows at a 35 days interval during a 154 day postpartum period. Figure 5.1 The effect of nutritional treatments differing after calving in energy levels and sources on the total number of ovarian follicles (mean ± SE) following an Ovsynch protocol recorded during a 21 ± 3 day period. Figure 6.1 The effect of nutritional treatments differing after calving in energy levels and sources on the cumulative proportion of pregnant primiparous cows (mean ± SE) recorded during a 154 day postpartum period. Figure 6.2 The effect of nutritional treatments differing after calving in energy levels and sources on the cumulative proportion of pregnant multiparous cows (mean ± SE) recorded during a 154 day postpartum period. Figure 7.1 A graphical representation of pasture quality parameters monitored during the postpartum observation period of the study. Figure 7.2 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on live weight (mean ± SE) recorded for primiparous dairy cows during an eight week prepartum to 17 week postpartum period. Figure 7.3 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on postpartum live weight loss (mean ± SE) recorded for primiparous dairy cows during a 17 week postpartum period. Figure 7.4 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on live weight (mean ± SE) recorded for multiparous dairy cows during an eight week prepartum to 17 week postpartum period. Figure 7.5 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on postpartum live weight loss (mean ± SE) recorded for multiparous dairy cows during a 17 week postpartum period. Figure 7.6 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on the milk yield (mean ± SE) recorded for primiparous dairy cows during a 17 week postpartum period. Figure 7.7 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on the milk yield (mean ± SE) recorded for multiparous dairy cows during a 17 week postpartum period. xix

20 LIST OF TABLES Table 3.1 The chemical composition of feeds for dairy cows used in the study. Table 3.2 Determined composition of the experimental control, HSLF and LSHF diets. Table 3.3 The effect of nutritional treatments differing after calving in energy levels and sources on the non-esterified fatty acid and β-hydroxybutyrate (mean ± SE) levels recorded for primiparous and multiparous dairy cows during a four week prepartum to 13 week postpartum period. Table 3.4 The effect of nutritional treatments differing after calving in energy levels and sources on the plasma urea (mean ± SE) levels recorded for primiparous and multiparous dairy cows during a four week prepartum to 13 week postpartum period. Table 3.5 The effect of nutritional treatments differing after calving in energy levels and sources on live weight traits (mean ± SE) recorded for primiparous and multiparous dairy cows during an eight week prepartum to 22 week postpartum period. Table 4.1 The effect of nutritional treatments differing after calving in energy levels and sources on the milk production and milk composition (mean ± SE) recorded for primiparous and multiparous dairy cows during a 22 week postpartum period. Table 5.1 The effect of nutritional treatments differing after calving in energy levels and sources on ovarian and follicular dimensions, follicular counts (mean ± SE) and proportions of Holstein cows ovulating and pregnant following an Ovsynch protocol recorded during a 21 ± 3 day period. Table 6.1 The effect of nutritional treatments differing after calving in energy levels and sources on LW changes and fertility traits (mean ± SE) recorded for primiparous and multiparous dairy cows during a 22 week postpartum period. Table 7.1 The chemical composition of feeds and estimated chemical composition of prepartum diets used in the study. Table 7.2 Determined composition of the experimental postpartum diet. Table 7.3 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on live weight traits (mean ± SE) recorded for primiparous and multiparous dairy cows from 8 week prepartum to 17 week postpartum period. Table 7.4 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on milk production and milk composition (mean ± SE) recorded for primiparous and multiparous dairy cows during a 17 week postpartum period. Table 7.5 The effect of prepartum nutritional treatments differing non-fibre carbohydrate and protein levels on LW changes and fertility traits (mean ± SE) recorded for primiparous and multiparous dairy cows during an eight week prepartum to 17 week postpartum period. xx

21 LIST OF ABBREVIATIONS AI: Artificial insemination AOAC: Association of Official Analytical Chemists ATP: Adenosine triphosphate BCS: Body condition score BHB: β-hydroxybutyrate Ca: Calcium CIDR: Controlled internal drug release CP: Crude protein d: Day D: Diameter DIM: Days in milk DM: Dry matter DMI: Dry matter intake DP: Dry period EB: Energy balance EE: Ether extract FCM: Fat corrected milk GIT: Gastro-intestinal tract GH: Growth hormone GLM: Generalised linear model GnRH: Gonadotropin releasing hormone IGF-I: Insulin-like growth factor-i I-R: Insulin receptor HSLF: High starch-low fat LH: Luteinizing hormone LSHF: Low starch-high fat LW: Live weight ME: Metabolizable energy MPS: Microbial protein synthesis MUN: Milk urea nitrogen MY: Milk yield N: Nitrogen xxi

22 NFC: Non-fibre carbohydrate NDF: Neutral detergent fibre NEB: Negative energy balance NEFA: Non-esterified fatty acids NE L : Net energy lactation NFC: Non-fibre carbohydrate NRC: National Research Council NS: Not significant P 4 : Progesterone P: Phosphor P: Probability PC: Pyruvate carboxylase PEPCKm: Phosphoenolpyruvate carboxykinase-mitochondrial RDP: Rumen degradable protein RNA: Ribonucleic acid PGF 2α : Prostaglandin-F 2α ptmr: Partial total mixed ration SAS: Statistical Analysis Systems S: Season of calving SD: Standard deviation SEM: Standard errors of mean ST: Interaction of season of calving and treatment t: Time T: Treatment TAG: Tri-acyl glycerol VFA: Volatile fatty acids Y: Year W: Week of observation WCDA: Western Cape Department of Agriculture WT: Interaction of week of observation and treatment xxii

23 Chapter 1 General introduction Over the past few decades, a significant increase in milk yield has been observed in dairy cows (Leroy et al., 2008a, b; Roche et al., 2011) because of an intense sire genetic selection, improved feeding of dairy cows while providing a better cow housing environment (Lucy, 2001; Thatcher et al, 2011). However, several studies have shown that these economically favourable improvements are associated with some negative physiological consequences, such as an increased incidence of metabolic (i.e. ketosis, fatty liver, lameness, etc.) and infectious diseases (i.e. mastitis, metritis), and a decline in fertility of dairy cows (Lucy, 2001, 2007; Butler, 2003; Walsh et al., 2011; Wathes, 2012). One of the major concerns of the physiological adaptation in mammals is that dry matter intake (DMI) does not meet the nutrient requirements during transition (e.g. at around calving), triggering a state of negative energy balance (NEB) (Butler, 2003; Ingvartsen, 2006; Roche et al., 2009). The state of NEB in dairy cows results in the mobilisation of body reserves (Rukkwamsuk et al., 1999) and the loss of LW (Jorristsma et al., 2003). In addition, it impairs the associated physiological and metabolic processes up to weeks postpartum, and has been identified as an underlying factor of poor reproductive performance of dairy cows (Butler & Smith, 1989; Garnsworthy & Webb, 1999; Butler, 2003; Jorristsma et al., 2003). Failures in fertility have transpired to delayed resumption of ovarian activity (Staples et al., 1990), reduced oestrous expression (Lopez et al., 2004), diminished viability and quality of oocyte (Lucy et al., 1991), poor conception rates and more days open (Reist et al., 2003; Resken et al., 2002). Currently, failure of dairy cows to conceive is considered a key reason for culling, threatening the longevity of cows, as well as the sustainability of the dairy industry (De Vries, 2006). During the postpartum period, different approaches to improve the reproductive performance in dairy cows have been put forward (Jorritsma et al. 2003; Lucy, 2007; Van Knegsel et al., 2007a, b; Garnsworthy et al., 2008a, b, c, d, 2009; Esposito et al., 2014). Most investigations found that improving the EB status during the transition period is of significance to ensure the subsequent well-being, milk production and fertility of dairy cows (Drackley et al., 2003). In this context, severe NEB during this periparturient period has been associated in the subsequent lactation to increased health problems (Duffield et al., 2009; Ospina et al., 2010a; McArt et al., 2012), decreased milk production and reduced reproductive success (Duffield et al., 2009; Ospina et al., 2010b, c). One suggested method, aiming for enhanced fertility while sustaining optimal milk yield, is to improve the energy density of the diet with non-fibre carbohydrates or fat (Staples et al., 1998; 1

24 Voigt et al., 2003; Gilmore et al., 2011). Dietary starch and/or fat increase the energy density of diets while reducing the severity of a NEB status (Van Knegsel et al., 2005). Dietary starch has been reported to improve insulin and glucose levels (Van Knegsel et al., 2007a, b; Garnsworthy et al., 2008b) and thus, have a positive influence on ovarian activity (Gong et al., 2002). However, a high level of dietary starch in the diet reduces the viability and quality of an oocyte and impairs the embryo developmental competence, thus leading to declining conception rates and a greater prevalence of embryo mortality (Armstrong et al., 2001; Leroy et al., 2008a, b). In contrast, dietary fat like calcium salts of palm fatty acids in the diet does not have inhibiting effects towards rumen microbial fermentation (Chalupa et al., 1986). Additionally, these fats were reported to enhance the growth and quality of follicles and also enhance progesterone (P 4 ) secretion, supporting an early embryo developmental competence (Gray et al., 2001; Wang et al., 2007). Several studies have investigated the possible benefits of energy sources during the prepartum period on postpartum energy balance (EB), milk production and fertility of dairy cows. However, results are limited and in some instances conflicting, with some studies reporting a positive effect on EB (Grum et al., 1996; Janovick & Drackley, 2010; Janovick et al., 2011; Damgaard et al., 2013), milk yield (Ingvartsen & Andersen, 2000; Cavestany et al., 2009a), milk composition (Cavestany et al., 2009b; Grum et al., 1996; Damgaard et al., 2013), and fertility success (Frajblat & Butler, 2003; Cavestany et al., 2009b), while others studies showed no effects on these parameters (McNamara et al., 2003; Agenäs et al., 2003; Burke et al., 2010; Mann et al., 2015). Feeding energy as starch and/or fat ad libitum to dry cows enhanced DMI, allowing them to over consume energy relative to their demands, compared with those being fed a diet with a lower energy density (Janovick & Drackley, 2010). This overconsumption of energy was reported to be detrimental to the cow rumen health and the liver function (Beever, 2006), resulting in a greater decline in DMI (Minor et al., 1998; Olsson et al., 1998). Furthermore, dietary fat affected the EB status negatively, as evidenced by increased plasma non-esterified fatty acids (NEFA) and β- hydroxybutyrate (BHB) levels and decreased plasma insulin levels during transition (Leroy et al., 2008b; Damgaard et al., 2013), which led to longer anoestrous periods (Giuliodori et al., 2011). In contrast, significant improvements of the cow health in early lactation have been reported in practice after feeding fibre-based diets containing > 400g/kg of NDF and low digestible energy levels on dry matter (DM) basis during the dry period (DP) (Drehmann, 2000; Beever, 2006; Jouany, 2006). Such diets were reported to optimise rumen microbial activity (Jouany, 2006), enhance metabolic status, reduce risks of ketosis and fatty liver syndrome (Janovick et al., 2011; 2

25 Vickers et al., 2013; Mann et al., 2015) and improve fertility outcomes in the subsequent lactation (Beever, 2006; Jouany, 2006). Against this background, studies investigating the effect of energy sources and levels before and after calving to mid- or late lactation on reproductive performances of dairy cows are limited. Also, no studies have been conducted in South Africa to investigate the prepartum and/or postpartum effect of energy nutrients on milk responses and reproductive performances of dairy cows. Appropriate nutrition programmes of dairy cows support not only the level of milk production but also ensure the rumen health, the restoration of body condition and energy stores between lactations, and the ability to calve at annually (Field & Taylor, 2012). The latter influences the culling rate, the size of the herd, the cow longevity and ultimately the financial return of lifetime milk production and the sustainability of dairy farming (De Vries, 2006; Leroy & De Kruif, 2006; Inchaisri et al. 2011). However, feed costs represent the largest single component (almost 70%) of the total input costs in dairy farming (Field & Taylor, 2012) and maintaining poor or non-fertile and/or lower milk producing cows in the dairy herd is obviously non-economical. Trials were conducted at the Elsenburg Research Farm of the Western Cape Department of Agriculture (WCDA, Ethical clearance Project AP/BR/D/CM31). These trials evaluated the energy levels and sources in the diet of dairy cows before and/or after calving to improve fertility of Holstein cows, while sustaining milk production. The first trial investigated the effect of nutritional treatments differing after calving in energy levels (low vs high) and sources (starch vs fat) on plasma metabolites and LW changes, milk production and milk composition, and reproductive performances of Holstein cows until 154 days postpartum. The treatments were: Control: ad libitum pasture + control concentrate (7 kg/day for both primiparous and multiparous cows). Glucogenic Treatment: ad libitum pasture + high starch-low fat (HSLF) concentrate (11.6 and 12.6 kg/day for primiparous and multiparous cows, respectively). The HSLF treatment was formulated to increase plasma insulin and glucose levels in order to reduce the extent of a NEB and improve an early resumption of oestrous activity and pregnancy rates in grazing cows. Glucogenic-lipogenic treatment: ad libitum pasture + high starch-low fat (HSLF) concentrates, followed from 61 days in milk (DIM) by low starch-high fat (LSHF) concentrate (11.6 and 12.6 kg/day of HSLF and LSHF concentrates for primiparous and multiparous cows, respectively). In addition to the HSLF effect during the first 60 DIM, the 3

26 LSHF concentrate was fed thereafter to decrease plasma insulin and increase plasma cholesterol in order to improve the ovulatory follicular size, the production of progesterone, and pregnancy rates in grazing cows. The second trial was simultaneously conducted with the first trial and investigated the effect of the same nutritional treatments (i.e. control, HSLF and HSLF-LSHF supplements) on the ovarian follicular dynamics and oestrous activity of grazing multiparous cows. The third trial studied the 30 day effect of prepartum nutritional treatments differing in non-fibre carbohydrate (NFC) and protein levels on LW changes, milk production and milk composition, and fertility traits of Holstein cows until 120 days postpartum. The treatments were: Control: Prepartum: ad libitum unchopped oat hay + prepartum concentrate. Postpartum: ad libitum pasture + postpartum concentrate. The prepartum concentrate levels were fed from 30 to 14 days prepartum at 3 kg/day for both heifers and dry cows, and from 13 days prepartum to calving at 5 kg/day and 6 kg/day for heifers and dry cows, respectively. The postpartum concentrate was fed at 7 kg/day for both primiparous and multiparous cows. Treatment : Prepartum: ad libitum partial total mixed ration (ptmr, consisting of 48% oat hay + 43% lucerne hay and 9% soybean oil cake meal) + prepartum concentrate. Postpartum: ad libitum pasture + postpartum concentrate. The prepartum concentrate levels were fed from 30 to 14 days prepartum at 3 kg/day for both heifers and dry cows, and from 13 days prepartum to calving at 5 kg/day and 6 kg/day for heifers and dry cows, respectively. The postpartum concentrate was fed at 7 kg/day for both primiparous and multiparous cows. The prepartum treatment was formulated to optimise rumen digestion and fermentation, through improved prepartum intake of energy and protein per kg on DM basis, to reduce the extent of a periparturient NEB, compared to the control group. The first hypothesis of this study was that manipulating the type and inclusion level of energy nutrients in the diet of dairy cows until 154 days postpartum can: influence the metabolite profiles and prevent a decrease in LW since the improved postpartum nutritional status [i.e. total nutrient intake and metabolizable energy (ME)] affects the extent of body fat mobilisation in the adipose tissues (Lafontan et al., 2009), the nitrogen (N) metabolism (Moharrery, 2004.) as well as the restoration of LW (Bauman & Currie 1980); 4

27 Improve milk yields and influence milk composition as made possible by an increase in the supply of milk precursors to the udder for milk synthesis (Van Knegsel et al., 2007a; Purushothaman et al., 2008; Hills et al., 2015); Enhance ovarian follicular dynamics and oestrous activity in an in vivo systems as in vitro systems do not seem to provide the complete in vivo ovarian condition (Laskowski et al., 2016); Improve fertility traits in dairy cows, as the improved nutritional status and the carry-over effects of the positive EB affect positively the reproductive axis and improve fertility outcomes (Butler, 2003). Contrary to lipogenic conditions, high insulinogenic conditions from high starch-based diets were reported in in vitro systems to affect negatively the viability and quality of the oocyte and later of the embryo (Armstrong et al., 2001; Leroy et al., 2008a, b). The second hypothesis of this study was that feeding fibre-based diets (> 400g/kg of NDF on DM basis) differing in NFC and protein levels during the DP can affect the energy and nutrient intake of periparturient dairy cows to subsequently minimise LW loss, influence milk production and milk composition, and improve the proportion of cows pregnant in the subsequent lactation as stated in some studies (Grum et al., 1996; Ingvartsen & Andersen, 2000; Cavestany et al., 2009a, b; Damgaard et al., 2013). The first aim of this thesis was to investigate in a pasture-based system the effect of nutritional treatments differing after calving in energy levels (low vs high) and sources (starch vs fat) on: 1. Plasma metabolites and LW changes; 2. Milk production and milk composition; 3. Ovarian follicular dynamics and oestrous activity; 4. Fertility traits of Holstein cows. The second aim of this thesis was to investigate the effect of two late prepartum fibre-based diets differing in levels (low vs high) of NFC and protein on: 5. Milk production and milk composition, LW changes and fertility traits of Holstein cows in the subsequent lactation in a pasture-based system. With the prepartum and/or postpartum application of the energy levels and sources in the trials, it is expected that a nutritional contribution of this study can be made not only to enhance plasma 5